Anti-laminin Antibodies Research Articles

Meniere's disease: Role of antibodies against basement membrane antigens

Recently, high levels of anti-collagen II (CII) antibodies have been found in a significant percentage of patients with Menière's disease (MD). An immune response against other collagens (C) present in the basal membranes has also been suggested by the presence of anti-CIV and anti-laminin antibodies in a number of instances of progressive systemic sclerosis (PSS) and Raynaud's phenomenon (RP). The aim of the present study was to measure the levels and frequency of anti-CI, -CII, -CIV, -CV, and anti-laminin antibodies by enzyme-linked immunosorbent assay (ELISA) in 45 patients affected by MD, 24 patients with PSS, 24 patients with RP, and 25 normal subjects. Immunoglobulin G (IgG) anti-CII antibodies were detected in 15.5% of MD sera, and 6.6% of MD sera were positive on CV, 11.1% were positive on laminin, 4.4% were positive on CI, and 4.4% were positive on CIV. A low incidence of anti-CI, -CII, -CIV, -CV, and antilaminin antibodies was found. Positive sera did not react on immunoblot with peptides obtained by cyanogen bromide (CNBr) digestion of CII. These results contrast with those of Yoo, et al.; differences in the washing procedure used and in the ethnic background of the patients may explain this divergence. Our results would seem to indicate that autoimmune responses do not play a role in the pathogenesis of MD.

An artificial esophagus consisting of cultured human esophageal epithelial cells, polyglycolic acid mesh, and collagen.

Previously, the authors successfully grafted collagen sheets covered with cultured human esophageal epithelial cells onto latissimus dorsi muscles of athymic mice. To make a tubular artificial esophagus they developed a polyglycolic acid mesh-collagen complex tube whose inner side was epithelialized in vitro by cultured human esophageal epithelial cells. Human esophageal epithelial cells were isolated from normal mucosa resected from specimens of esophageal cancer patients and cultured on the surface of the collagen gel in which polyglycolic acid mesh was embedded. After the culture reached confluence, polyglycolic acid mesh was sutured to create a tubular form lined with cultured cell layers (restructured mucosal tube). Such tubes were wrapped in the latissimus dorsi muscle flaps of athymic rats using a fibrin glue. The animals were killed 4, 8, 20, and 28 days after grafting and the grafted tubes were studied histologically. Up to 28 days after grafting, the luminal structure was maintained without structure. Eight days after grafting, rat fibroblasts infiltrated from the muscle and neovascularization appeared in the collagen layer. The grafted epithelium remained healthy and grew to 15 cell layers by 20 days, much like normal human esophageal epithelium. Staining by anti-laminin monoclonal antibody confirmed a basement membrane.

Cell surface laminin-like substances and laminin-related carbohydrates of rat ascites hepatoma AH7974 and its variants with different lung-colonizing potential

Rat ascites hepatoma AH7974 cells strongly expressed antilaminin antibody-reactive substances (laminin-like substances) and Griffonia simplicifolia isolectin B4 (GS)-reactive carbohydrate (alpha-D-galactose; alpha-Gal) on their cell surface. The alpha-Gal expression was not apparently influenced by the pretreatment of cells with methanol. The cell membrane laminin-like substances has approximate molecular weights of 150, 62 and 56 kDa in denaturating reducing conditions, of which the 62 and 56 kDa bands were stained with GS. The cell membrane molecules bearing alpha-Gal were 62 and 56 kDa and were stained with antilaminin antibody. Therefore, the major molecules bearing alpha-Gal residues of AH7974 cell membrane are considered to be laminin-like substances. To determine the role of the substances in metastasis, we selected four cell lines (74AD, 74AD-f, 74FL, 74FL-a) from AH7974 in culture. 74AD and 74FL-a are adherent lines and 74AD-f and 74FL are floating lines. All of these cell lines strongly expressed laminin-like substances, but a marked difference was found in expression of alpha-Gal, which was most strongly expressed by 74FL, followed by 74AD, and rarely by 74AD-f and 74FL-a; the staining intensity was positively correlated with their experimental lung-colonizing potential. Cell membrane laminin-like substances were 200, 97, 62, 56 and 46 kDa and among them 62 and 56 kDa molecules were glycosylated with alpha-Gal. The pretreatment of 74FL cells with antilaminin antibody or with human type A serum (containing natural antibody to alpha-Gal epitope) depressed remarkably the lung-colonizing potential of the cells. These results suggest that the expression of 62 and 56 kDa laminin-like substances with alpha-Gal residues on tumor cell surfaces is one of the determinants associated with lung-colonizing potential of these cells.

An acidic component of the heterogeneous Tc-85 protein family from the surface of Trypanosoma cruzi is a laminin binding glycoprotein

Successful infection of mammalian host by trypomastigotes of Trypanosoma cruzi is a complex event, involving host receptors and parasite ligands. Interaction of the trypomastigote stage with laminin, a component of specialized extracellular matrices, as basement membranes, is studied in this report. Binding of 125I-laminin to trypomastigotes is specific and 2−5 × 10 3 laminin binding sites were calculated to be present on the surface of live trypomastigotes. Antilaminin antibodies were able to inhibit the invasion of cultured cells by trypomastigotes (75-62%), suggesting that laminin may be involved in the adhesion of the parasite to host cells. By affinity chromatography, an 85-kDa glycoprotein was isolated (laminin binding glycoprotein, LBG) from trypomastigote lysates, but not from epimastigote lysates. It is suggested that at least fragment E8 (but not E1′) from laminin could be involved in the reaction which is independent of the carbohydrate moieties from both ligand and receptor, as suggested by glycosidase or tunicamycin treatments. It is also shown that LBG is an acidic component of the polymorphic Tc-85 protein family, a trypomastigote-specific surface membrane glycoprotein which contains several polypeptides recognized by the monoclonal antibody H1A10, and previously related with the invasion process of the parasite.

Immunocytochemical and functional characterization of an immortalized type 1 astrocytic cell line

The DI TNC 1 cell line has been derived from cultures of rat brain astrocytes targeted oncogenesis. Cultured astrocytes are known to promote neurite outgrowth via the production of adhesion molecules found either on the cell surface or in the extracellular matrix. We sought to investigate whether DI TNC 1 cells retained the ability to produce such neurite-inducing molecules, and promote neurite growth. We found by immunofluorescence that DI TNC 1 cells expressed laminin, N-CAM and 1A1. The latter is a cell adhesion molecule that is expressed exclusively on astrocytes of the type 1 lineage. In vitro neurite outgrowth assays were also used to assess the functional properties of these cells. Monolayers of DI TNC 1 cells were almost as effective a substrate as monolayers of astrocytes purified from the neonatal rat brain in their ability to support neurite outgrowth. In addition, PC12 cells grown on extracellular matrix derived from either DI TNC 1 cells or neonatal astrocytes displayed significantly more neurite growth than cells plated on plastic. This effect was partially inhibited by preincubation of the extracellular matrix with anti-laminin antibodies. Taken together, these results suggest that the immortalized DI TNC 1 cells show many similarities to neonatal astrocytes. Given the heterogeneity of cultured astrocytes, this homogeneous cell line may prove to be particularly useful for future investigations on interactions between glia and neurons.

Anti-laminin autoantibodies in collagen vascular diseases: the use of adequate controls in studies of autoimmune responses to laminin.

To determine the significance of anti-laminin antibodies in patients with collagen vascular diseases using a large control population of normal individuals. Anti-laminin antibodies of IgG isotypes were determined using an ELISA assay in a population consisting of 73 patients with systemic sclerosis, 10 with urticarial vasculitis, five with leukocytoclastic vasculitis, 13 with giant cell arteritis, and eight with dermatomyositis. Sera from 134 healthy individuals served as controls. Only eight sera in the systemic sclerosis group, one in the leukocytoclastic group and one in the giant cell arteritis group were abnormal. No other sera were abnormal. These data contradict previous studies using smaller numbers of controls. The necessity for an adequate control population to define abnormal when comparing immune responses among groups is frequently under-emphasised in rheumatological studies.

Immunohistochemistry of parasitic subepidermal vesiculobullous disease in American badgers (Taxidea taxus).

Some populations of free-ranging American badgers (Taxidea taxus) develop a distinctive seasonal dermatitis due to the subcutaneous filariid Filaria taxideae. Subepidermal vesicles that contain filarial ova develop in thinly haired skin of the inguinal area, proximal thigh, and ventral abdomen. The purpose of this study was to establish by immunohistochemistry whether basement membrane components colocalized with the roof or floor of vesicles and to confirm that filarial ova occur in intradermal vessels. Samples of skin with characteristic F. taxideae-induced subepidermal vesicles were collected from 10 adult male (n = 8) and female (n = 2) badgers. Samples were fixed in formalin for 1-4 days and processed routinely into paraffin wax. Immunohistochemical staining for basement membrane was attempted with anti-collagen IV antibodies (AM168-5M, AR079-5R, AB748) and antilaminin antibodies (MA078-5C, AR078-5R, L-9393). Optimal results in skin from badgers were obtained using a biotin-streptavidin technique and AR079-5R (anti-human collagen IV) and AR078-5R (anti-murine laminin). There was positive staining of the floor of vesicles in 5 of 6 badgers tested with antibodies to laminin and collagen IV. In 5/10 badgers, filarial ova and first stage F. taxideae larvae were found in dilated vascular channels of the upper dermis, and these vessels stained positively for factor VIII-related antigen. The results suggest that F. taxideae-induced subepidermal separation occurs consistently in the lamina lucida portion of the basal lamina and that filarial ova occur in dermal vessels.

Movement of embryonic chick sympathetic neurons on laminin in vitro is preceded by neurite extension.

Chick sympathetic neurons (E-9) are capable of moving on a laminin substrate but not on more adhesive substrates in vitro. The effect of laminin is dose-dependent and reduced by the addition of anti-laminin antibodies, whereas soluble laminin does not stimulate movement. The onset of neuronal movement is preceded by, and highly correlated with, the onset of neurite formation. The addition of 1,2 dioctanoyl-sn-glycerol (DAG), a stimulator or protein kinase C that has been shown to inhibit neurite outgrowth, was found to delay both process formation and neuronal movement but did not affect the correlation between these two measures. These results support the conclusion that laminin stimulates primary neuronal movement in vitro and suggest that the mechanism underlying movement involves process formation followed by "towing" of the cell body by the advancing process. The similarities of this in vitro behavior to that observed in vivo suggest that similar mechanisms may underlie neuronal movement in the developing nervous system as suggested by Morest (Z Anat Entwicklungsgesch 130:265-305, 1970) and Liesi (EMBO J 4:1163-1170, 1985; Exp Neurol 117:103-113, 1992).

Cranin interacts specifically with the sulfatide-binding domain of laminin.

Cranin is a 120 kDa integral membrane glycoprotein which binds laminin under conditions of physiologic ionic strength in a calcium-dependent manner. Here, binding of cranin to laminin has been characterized using both ligand-blotting assays and laminin affinity bead assays. Binding was specifically inhibited by anti-laminin antibodies against the A chain terminal domain G, but not by several other region-specific antibodies. Dextran sulfate, fucoidin, and sulfatide were potent inhibitors of binding (50% inhibition at 0.03, 0.5, and 1.7 micrograms/ml, respectively); heparin was a weaker inhibitor (50% inhibition approximately 5 micrograms/ml), and mannan and chondroitin sulfate were not inhibitory at 100 micrograms/ml. Binding was not inhibited by lactose or the A chain peptide PA22-2. The mobility of the broad, fuzzy cranin band was shifted after digestion with neuraminidase, N-glycanase, and O-glycanase, though none of these treatments decreased band heterogeneity nor destroyed the ability to bind laminin. Cranin bound to Jacalin lectin, which recognizes the Gal beta 1-3GalNAc linkage expressed in certain classes of mucins. These findings indicate that cranin binds at or near the high affinity sulfatide-binding site previously mapped to the E3 domain of laminin, which is known to exhibit bioactivity for neural cells. In view of the extremely low abundance of cranin in brain membranes (approximately 0.005%), its avid laminin-binding activity is remarkable, and strongly suggests that cranin may play a physiologic role in regulating specific neural cell interactions.

Immunohistochemical localization of laminin in the hearts of patients with chronic chagasic cardiomyopathy: Relationship to thickening of basement membranes

Histologic, ultrastructural, immunohistochemical studies were made of endomyocardial biopsy specimens from 10 patients with chronic chagasic cardiomyopathy. Histologic and electron microscopic observation disclosed marked thickening of the basement membranes of the myocytes, endothelial cells, and vascular smooth muscle cells in all patients. Light (peroxidase-labeled antibodies) and electron (gold-conjugated antibody) microscopic immunohistochemical methods revealed a positive reaction for laminin in these thickened basement membranes. This thickening of basement membranes may develop as a consequence of: (1) an immunologic reaction that is triggered by the presence of a laminin-like molecule on the surfaces of Trypanosoma cruzi amastigotes and trypomastigotes; (2) an immunologic response to direct injury of basement membranes causing some of their components to become antigenic; (3) myocardial fibrosis, with synthesis of new connective tissue components; and (4) a combination of the preceding factors. The relationship of these changes to antilaminin antibodies requires clarification.

Immunocytochemistry of Skeletal Muscle Basal Lamina Grafts in Nerve Regeneration

The influence on nerve regeneration of the extracellular matrix glycoprotein laminin was studied after sciatic nerve transection in 90 outbred Sprague-Dawley rats. Nerve regeneration through basal lamina grafts was comparable with regeneration through traditional nerve grafts across gaps up to 2.0 cm in length. True axonal regeneration rather than axonal branching was demonstrated by retrograde horseradish peroxidase labeling of nerve cables. Pretreatment of basal lamina grafts with antilaminin antibodies reduced the total number of regenerated axons by 90 percent with a significant decrease of nerve conduction velocity and a significant impairment of walking track patterns. The basement membrane glycoprotein laminin serves a critical role in the regeneration of peripheral nerves through basal lamina grafts.

Molecular analysis of spontaneous nephrotropic anti-laminin antibodies in an autoimmune MRL-lpr/lpr mouse.

To explore the genetic relationship between anti-laminin and anti-DNA autoantibodies (autoAb), VH gene and gene family expression were determined among autoAb derived from an individual 6-mo-old MRL-lpr/lpr mouse. Whereas 85% of the anti-DNA Ig were identified by one of two VH family probes, 7183 and VHJ558, none of the anti-laminin antibodies (Ab) examined were recognized by these probes. Subsequent V region sequence analysis of three of the anti-laminin Ab revealed that they in fact utilized a J558 VH gene (VH50). Furthermore, FR2 and CDR2 oligonucleotide probes complementary to VH50 recognized multiple anti-laminin Ab by Northern blot analysis; the FR2 probe recognized two control anti-DNA Ab, but neither probe recognized anti-DNA Ab from the same mouse. Polymerase chain reaction amplification of MRL-lpr/lpr genomic liver DNA using primers generated from VH50 and Vk50 sequences indicated that all three anti-laminin Ig have a single replacement mutation in both their VH and Vk genes. Search of the nucleic acid databases revealed that both germline VH and Vk genes are expressed unmutated by murine lupus anti-dsDNA autoAb, previously sequenced in other laboratories. Sequence comparisons suggest that differences in anti-DNA and anti-laminin reactivity may be dependent upon somatically generated differences in the CDR3 regions of the H and L chains. The results indicate that lupus anti-laminin Ab can arise from distinct B cell populations but express the same unmutated germline V region genes as lupus anti-dsDNA autoAb. They further raise the possibility that these distinct B cell populations may be activated and expanded either: independently, by distinct Ig receptor ligands such as the Ag, laminin and DNA; or simultaneously, by a common ligand such as an anti-Id recognizing a common V region epitope.

Role of laminin autoantibodies on the embryo toxicity of sera from mercuric chloride treated brown Norway rats

In previous studies, antilaminin antibodies were found to be toxic to cultured rat embryos. In order to extend these studies, Brown Norway rats were treated with mercuric chloride, which led to the production of laminin autoantibodies. Sera samples from Brown Norway rats treated with mercuric chloride were found to be teratogenic as well as lethal to cultured rat embryos. This embryotoxicity was not associated with sera mercury levels, but was related to the levels of antilaminin antibodies in sera. Affinity purified laminin antibodies from these mercuric chloride treated Brown Norway rats, when added to control sera, were found to be teratogenic but not lethal. These antibodies were found to bind to the laminin sequences IKVAV (A chain) and YIGSR (B1 chain), but not RGD (A chain) or YD (B1 chain). These observations suggested the possibility that an environmental pollutant such as mercury could cause the formation of embryotoxic autoantibodies that could persist in the body as embryotoxic factors for extended periods of time.

Propolypeptide of von Willebrand factor serves as a substrate for factor XIIIa and is cross-linked to laminin

The propolypeptide of von Willebrand factor (pp-vWF) was found to serve as a substrate for Factor XIIIa (FXIIIa). FXIIIa catalyzed polymerization of pp-vWF as well as the incorporation of monodansylcadaverine and [14C]putrescine into pp-vWF in a Ca(2+)- and time-dependent manner. The amount of putrescine incorporated into pp-vWF was increased as a function of putrescine concentration and reached a maximum of 3 mol/mol of pp-vWF, indicating that there are at least 3 glutamine residues in the pp-vWF molecule responsible for the reaction. 125I-pp-vWF was incubated with FXIIIa in the presence of various adhesive glycoproteins. Laminin, among the proteins tested, specifically formed a large molecular weight complex with 125I-pp-vWF in a Ca(2+)- and time-dependent manner. It was also found that FXIIIa catalyzed polymerization of laminin as well as the incorporation of monodansylca-daverine into laminin. The high molecular weight complexes of 125I-pp-vWF (or 125I-laminin) formed in the presence of laminin (or pp-vWF) were immunoprecipitated with anti-laminin (or anti-pp-vWF) antibodies. Taking all the data together it can be concluded that both pp-vWF and laminin have glutamine and lysine residues responsive to FXIIIa and make copolymers by virtue of FXIIIa.

Laminin cleavage by activated human neutrophils yields proteolytic fragments with selective migratory properties.

We studied the interactions between human neutrophils, as well as the purified human neutrophil serine proteases elastase (HNE) and cathepsin G (HNCG), and laminin. Our results show that intact laminin and two proteolytic fragments generated by HNE bind to neutrophils and stimulate cell migration. Domain-specific antilaminin monoclonal antibodies, rotary shadowing electron microscopy, and Western blotting mapped the two promigratory fragments on the laminin cross to the apical three-armed region and long arm, respectively. In contrast, a fragment derived from the terminal ends of short arms neither bound to neutrophils nor stimulated migration. When neutrophils embedded in a reconstituted basement membrane gel were activated with phorbol myristate acetate, several stable, proteolytic laminin fragments were released into supernatants. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting showed that these fragments appeared identical to those generated after digestion of soluble laminin with HNE and HNCG. Furthermore, release of laminin fragments by embedded neutrophils was inhibited by diisopropyl fluorophosphate, and duplicated by incubating the basement membrane gel with purified HNE and HNCG. Our findings therefore suggest that neutrophils, through release of HNE and HNCG, are capable of digesting basement membrane laminin in vivo. In addition, the release of laminin fragments from damaged basement membranes may promote neutrophil migration and thereby accelerate inflammatory processes.

Immunohistochemistry of the human ductus epididymis.

Our objective was to characterize epithelial cells, lamina propria, and sites of estrogen coupling in the caput, corpus, and cauda regions of the human epididymis using antibodies to cytokeratin types; epithelial membrane antigen; laminin; type IV collagen; vimentin; desmin-, and estradiol-receptor-related protein; and immuno-histochemical techniques. Principal cells immunostain by both AE1/AE3 antibodies (keratins 1-8, 10, 13-15, and 19) and anti-pan-keratin antibodies (keratin 5, 6, and 8). Immunoreactions to both anti-keratin antibodies increase from the caput to the cauda epididymis. The principal cells only immunostained by anti-keratin 19 antibodies in the cauda and showed no reaction to keratins 10 and 11. Basal cells and apical cells immunoreact to anti-AE1/AE3, antipankeratin, and antikeratin 19 antibodies, but not to antikeratin 10 and 11 antibodies, in all three epididymal regions. The principal cells immunoreact with epithelial membrane antigen antibodies in the stereocilia and subjacent cytoplasm. This immunostaining decreased from the caput to the cauda. Antivimentin antibodies stained the apical cytoplasm of principal cells and limited areas of both principal cells and basal cells. This immunoreaction decreased from the caput to cauda. Apical cells immunostained in the three regions. Immunoreaction to ER-D5 was moderate in the principal cells, basal cells, apical cells, and muscular coat cells in the cauda. The apical cells immunostained in the three regions. Antilaminin antibodies stained the epithelial basement membrane in the three regions. Type IV collagen was detected in the basement membrane as well as around the muscular coat cells in the three regions. Immunoreaction to desmin was intense in the muscular coat cells in the three regions.(ABSTRACT TRUNCATED AT 250 WORDS)

Clinical significance of laminin deposition and T-cell infiltration in oral cancer.

Biopsied specimens from 55 patients with squamous cell carcinoma (SCC) of the oral cavity were examined immunohistopathologically as to the clinical significance of basement membrane (BM) deposition and T-cell infiltration at the tumor-stromal border using monoclonal anti-laminin and anti-CD3 antibodies. According to the immunoreactivity, all specimens could be divided into three groups: group A, a continuous linear pattern of positive staining for BM around tumor nests; group B, an alteration of BM deposition around tumor nests with T cell infiltration into those tumor nests; and group C, an alteration of BM deposition around tumor nests without T cell infiltration into those tumor nests. These groups were correlated with clinical manifestations, such as tumor size, tumor regression rate with induction chemotherapy, and regional lymph node metastatic rate. In these groups, tumors classified as group C showed a trend toward resistance to chemotherapy and high metastatic characteristics. Tumors classified as group B, which showed the same alteration of BM deposition as a result of T cell infiltration into the tumor nests, showed a sufficient tumor regression rate with chemotherapy. The visualization of the staining for BM laminin and T cells in oral SCC appeared not only to increase our understanding of the biologic and clinical behavior of individual tumors, but could be a prognostic indicator.

Immunosuppressive therapy of organ-specific nephritic autoimmune diseases with 15-deoxyspergualin.

According to its immunopharmacological profile, 15-deoxyspergualin (15-DOS) has been investigated as to its disease-modifying activity on HgCl2-induced glomerulonephritis (GN) and on tubulointerstitial nephritis (TIN) in Brown-Norway rats. Both models are induced autoimmune disorders in which afflicted animals display high levels of serum autoantibodies directed against the glomerular or tubular basement membrane (GBM or TBM), respectively. The diseases are manifested by high serum creatinine and urea levels with severe proteinuria. In the model of HgCl2-GN, administration of 15-DOS clearly led to a reduction of proteinuria and decreased the amount of rat IgG attached to the GBM. Furthermore, a therapeutic effect could be demonstrated when 15-DOS was given after the appearance of clinical symptoms. Not only urine-protein values but also anti-laminin antibodies returned to normal levels. Also in the experimental TIN-model, 15-DOS, either given during the induction phase, or even late in the onset of the disease, strongly prevented the proteinuria of this autoimmune disease and inhibited the formation of autoantibodies to TBN.

Autoantibodies in the sera of Trypanosoma cruzi-infected individuals with or without clinical Chagas disease.

Variations in the levels and the specificities of autoantibodies directed against a panel of antigens (cytoskeleton proteins, DNA, laminin) were analyzed in the sera from two groups of humans infected with Trypanosoma cruzi. One group was constituted of apparently healthy blood donors (BD) and the other of patients with clinically confirmed Chagas disease (CCH). In both infected groups, a high proportion but not all sera exhibited dramatic enhancement of IgM and IgG autoantibodies directed against all antigens tested. Sera positive for IgG autoantibodies were generally found more frequently in the CCH than in the BD group, except for anti-actin antibodies more often present in BD sera. Anti-laminin IgG antibodies were present in a similar number of individuals in both groups. Although the titers of anti-laminin IgG antibodies were in general higher in CCH, their dissociation constants were in the same range (7 x 10(-8) - 10(-7) M) in both groups. IgG autoantibodies were demonstrated to be polyreactive with laminin and other self antigens as well. Circulating immune complexes were present in sera from both groups and the activity of the antibodies dissociated from these complexes was directed against all the antigens of the panel. Although the IgE concentration was significantly enhanced in several subjects from both groups, the incidence of positive sera was higher in the CCH (60%) than in the BD (39%) group. Our results demonstrate that autoantibodies with the characteristics of natural autoantibodies are found in both T. cruzi-infected apparently healthy individuals and patients.

Evidence for a role of antilaminin-producing B cell clones that escape tolerance in the pathogenesis of HgCl<sub>2</sub>-induced membranous glomerulopathy

In Brown-Norway rats HgCl2 induces an autoimmune disease due to a T-dependent B cell polyclonal activation. This disease is marked by the production of numerous antibodies including antiglomerular basement membrane (GBM) antibodies. Rats exhibit a biphasic glomerulopathy with heavy proteinuria. Initially anti-GBM antibodies are found linearly deposited; they precede the appearance of membranous glomerulopathy. Rats recover spontaneously even if HgCl2 injections are pursued, but mechanisms at play are unclear. We have assessed the effects of transplanting the spleen from a BN rat, either at the acme of the disease or at the time of convalescence, into naive BN rats, some of which were then injected with HgCl2. Transplantation of a spleen from HgCl2-injected rats at the acme of the disease dramatically protects BN rats from all the manifestations of the mercury disease. BN rats transplanted with a spleen from HgCl2-injected rats at the time of convalescence only exhibited a typical membranous glomerulopathy with heavy proteinuria but without circulating anti-GBM antibodies. Antilaminin antibodies were eluted from the glomeruli. This study shows that spleen cells from HgCl2-injected rats are able to confer tolerance to HgCl2-induced autoimmunity. It also shows that some B cell clones escape this tolerance. Finally, this study strongly suggests that membranous glomerulopathy, responsible for proteinuria in this model, is related to the presence of antilaminin antibodies.